Comparative Analysis of Root Canal Filling Debris and Smear Layer Removal Efficacy Using Various Root Canal Activation Systems during Endodontic Retreatment.

The complete removal of obturation material can be a challenge in nonsurgical root canal retreatment. The insufficient removal of obturation material is a reason for root canal retreatment failure. The purpose of this study was to assess the efficacy of different final root canal irrigation activation methods in removing debris and smear layers in the apical and middle portions of root canals during retreatment. Sixty-six distal roots of freshly extracted molars were randomly divided into six groups: (1) primary root canal treatment with no obturation (negative control); (2) retreatment with only conventional instrumentation and irrigation (positive control); (3) retreatment with additional ultrasonic irrigation using the Piezon Master 700; (4) ultrasonic irrigation with the ENDOSONIC Blue; (5) sonic irrigation with the EDDY; and (6) multisonic irrigation with the GentleWave system. Roots were split and prepared for scanning electron microscopic (SEM) evaluation. Acquired images were assessed to quantify the amount of debris and smear remaining. Among the treatment groups, Group 6 had a significantly lower debris score than Group 2 (positive control) in both the middle and apical regions (p = 0.004, p = 0.012). All treatment groups showed significantly lower smear scores than Group 2 in the middle and apical regions (p < 0.05). The GentleWave multisonic System showed a more optimal cleaning efficacy of the root canal debris but did not differ significantly with the tested passive ultrasonic or sonic irrigation method.

Kaposi's sarcoma-associated herpesvirus ORF18 and ORF30 are essential for late gene expression during lytic replication.

UNLABELLED: Kaposi's sarcoma-associated herpesvirus (KSHV) is associated with several human malignances. As saliva is likely the major vehicle for KSHV transmission, we studied in vitro KSHV infection of oral epithelial cells. Through infection of two types of oral epithelial cells, normal human oral keratinocytes (NHOKs) and papilloma-immortalized human oral keratinocyte (HOK16B) cells, we found that KSHV can undergo robust lytic replication in oral epithelial cells. By employing de novo lytic infection of HOK16B cells, we studied the functions of two previously uncharacterized genes, ORF18 and ORF30, during the KSHV lytic cycle. For this purpose, an ORF18-deficient virus and an ORF30-deficient virus were generated using a mutagenesis strategy based on bacterial artificial chromosome (BAC) technology. We found that neither ORF18 nor ORF30 is required for immediately early or early gene expression or viral DNA replication, but each is essential for late gene expression during both de novo lytic replication and reactivation. This critical role of ORF18 and ORF30 in late gene expression was also observed during KSHV reactivation. In addition, global analysis of viral transcripts by RNA sequencing indicated that ORF18 and ORF30 control the same set of viral genes. Therefore, we suggest that these two viral ORFs are involved in the same mechanism or pathway that coregulates the viral late genes as a group. IMPORTANCE: While KSHV can infect multiple cell types in vitro, only a few can support a full lytic replication cycle with progeny virions produced. Consequently, KSHV lytic replication is mostly studied through reactivation, which requires chemicals to induce the lytic cycle or overexpression of the viral transcriptional activator, RTA. In this study, we present a robust de novo lytic infection system based on oral epithelial cells. Using this system, we demonstrate the role of two viral ORFs, ORF18 and ORF30, in regulating viral gene expression during KSHV lytic replication. As the major route of KSHV transmission is thought to be via saliva, this new KSHV lytic replication system will have important utility in the field.

Comparative Analysis of Biofilm Removal Efficacy by Multisonic Ultracleaning System and Passive Ultrasonic Activation.

The purpose of this study was to compare disinfection and the biofilm removal efficacy of the GentleWave System (Sonendo, Inc., Laguna Hills, CA, USA) with passive ultrasonic activation method. Forty-seven freshly extracted human molars were inoculated with Enterococcus faecalis and cultured for five weeks to establish biofilm. Eight molars were tested for confirmation of infection. Four of the eight teeth were not inoculated in order to provide a negative control. The remaining 39 inoculated molars were randomly separated into three treatment groups (n = 13 per group): Group 1-no treatment, Group 2-conventional rotary instrumentation and passive ultrasonic activation, and Group 3-minimal instrumentation and the GentleWave System treatment. Roots were subsequently prepared per standard histological tissue processing procedures. Modified Brown and Brenn stained sections and Hematoxylin and Eosin stained sections were visualized at 4× and 13.5× magnification using a stereomicroscope. The sections were scored and blindly analyzed by two independent evaluators, including a histopathologist, to evaluate the presence of biofilm on canal wall. A significant difference was found between Group 2 and Group 3 in both apical and middle regions (p = 0.001) of the mesial roots of mandibular molars and mesiobuccal roots of maxillary molars. Group 3 revealed significantly less biofilm than the controls (p = 0.003). The GentleWave System demonstrated significantly greater reduction in biofilm within the mesial roots of mandibular molars and mesiobuccal roots of maxillary molars than those treated with conventional rotary instrumentation and passive ultrasonic activation protocol.

In vitro characterization of human dental pulp stem cells isolated by three different methods.

OBJECTIVES: In this study, we characterized human dental pulp cells (HDPCs) obtained by different culture methods to establish the most suitable methodology for dental tissue engineering and regenerative endodontic applications. MATERIALS AND METHODS: HDPCs were isolated by the outgrowth method (HDPCs-OG), the enzymatic digestion method (collagenase/dispase/trypsin, HDPCs-ED), or the combination of both methods (HDPCs-Combined). The expression of mesenchymal stem cell markers (CD105, CD90, and CD73) was investigated. In vitro differentiation capacities of HDPCs into adipogenic, osteogenic, and chondrogenic lineages were compared. Differentiation markers were analyzed by quantitative reverse-transcription polymerase chain reaction (RT-PCR) and western blotting. RESULTS: Our data indicated that whole HDPCs-ED, HPDCs-OG, and HDPCs-Combined could be differentiated into adipogenic, chrondrogenic, and osteogenic cell types. However, we found that the methods for isolating and culturing HDPCs influence the differentiation capacities of cells. HDPCs-OG and HDPCs-ED were preferably differentiated into adipogenic and osteogenic cells, respectively. Differentiation markers shown by RT-PCR and western blotting analysis were mostly upregulated in the treated groups compared with the control groups. CONCLUSIONS: Our findings confirmed that cell populations formed by two different culture methods and the combined culture method exhibited different properties. The results of this study could provide an insight into regenerative endodontic treatment using HDPCs.